Chronic obstructive pulmonary disease (COPD) is the third leading cause of death worldwide. In the USA alone, COPD induces $30-$50 billion in health care expenses each year. Despite this, a cure for COPD has yet to be found. Our group has recently shown that cigarette smoke (CS) exposure, a major cause of COPD, leads to rapid internalisation of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel and the formation of intracellular CFTR aggregates1. Since the internalisation of CFTR may contribute to mucus dehydration seen in COPD patients and particularly in chronic bronchitis (CB), we set out to elucidate the cellular mechanisms underlying this internalisation. To investigate if CS exposure induces aggregation in the plasma membrane prior to internalisation, we used GFP- and RFP-tagged CFTR constructs in Förster Resonance Energy Transfer (FRET)-based experiments. HEK293T Ccells expressing both GFP-tagged and RFP-tagged CFTR were exposed to freshly prepared gaseous phase of cigarette smoke or air control and incubated at 37oC for 15, 30 or 60 min before being fixed with 4% paraformaldehyde. FRET levels after 15, 30 and 60 min CS exposure (15.0 ± 1.2%, n = 48 cells; 16.2 ± 1.5%, n = 33; and 10.8 ± 1.4%, n = 34 cells, respectively; mean ± SEM) remained unchanged compared to air exposed cells at the same time points (17.4 ± 1.6%, n = 52; 12.7 ± 1.1%, n = 41; 12.3 ± 0.8%, n = 60 respectively; mean ± SEM) suggesting that CFTR aggregates intracellularly post-CS exposure rather than at the membrane. As an additional control we used Anoctamin 1 (Ano1), the protein underlying calcium-activated chloride secretion. Consistent with previous studies showing that CaCC secretion is not affected by CS1, we also saw no change in FRET levels after CS exposure (baseline FRET was 17.3 ± 1.2%, n = 32; mean ± SEM). Since CFTR’s C-terminus is known to regulate its trafficking, we set out to elucidate whether this region of CFTR was responsible for CS-induced internalisation. Confocal imaging of HEK293T cells showed that truncation of GFP-tagged CFTR to delete the C-terminal PDZ-binding motif (L1254X) or nucleotide binding domain 2 (NBD2) (K1174X) had no effect on the intracellular accumulation of CFTR post-CS. Thus we conclude that despite the established involvement of the C-terminus in normal CFTR trafficking, neither CFTR’s PDZ-binding domain nor NBD2 have a role in CS-induced CFTR internalisation. Further mutations are to be carried out in an effort to understand which domain(s) of CFTR are involved in the response to CS exposure.